Department of Pathophysiology, School of the Basic Medicine, Xuzhou Medical University, Xuzhou, 221004, China.
School of Nursing, Xuzhou Medical University, Xuzhou, 221004, China.
J Mol Neurosci. 2017 Dec;63(3-4):403-411. doi: 10.1007/s12031-017-0994-x. Epub 2017 Oct 29.
Parkinson's disease (PD) mainly results from the progressive loss of dopaminergic (DA) neurons in the substantia nigra pars compacta (SNpc), and the exact underlying mechanisms of the loss of DA neurons in PD remains largely unclear. The results of our previous work showed that let-7d was significantly downregulated in a 6-OHDA-induced cellular model of PD. However, the exact effect of let-7d on DA neural cells was unclear. In MN9D dopaminergic neuronal cells, we used a let-7d mimic and inhibitor to upregulate and downregulate the expression of let-7d, respectively, a cell counting kit to assess cell viability, and a TUNEL staining assay and flow cytometry to examine the cell death rate, and we found that let-7d could negatively regulate 6-OHDA-induced cell injury. Then, we verified that caspase-3 was a target gene of let-7d by using a dual-luciferase reporter system. Furthermore, using caspase-3 siRNA and a caspase-3-overexpression vector (without the 3'UTR) to respectively inhibit and increase the expression of caspase-3, we found that caspase-3 siRNA could reverse the cell injury induced by the let-7d inhibitor and that caspase-3 overexpression could reverse the protective effects of the let-7d mimic on 6-OHDA-induced cell injury. Taken together, these findings strongly suggest that let-7d plays an important role in DA neuronal cell injury and that the effects of let-7d are, at least in part, via the suppression of caspase-3 expression.
帕金森病（PD）主要是由于黑质致密部（SNpc）中的多巴胺能（DA）神经元进行性丧失引起的，而 PD 中 DA 神经元丧失的确切潜在机制仍很大程度上不清楚。我们之前的工作结果表明，let-7d 在 6-OHDA 诱导的 PD 细胞模型中显著下调。然而，let-7d 对 DA 神经细胞的确切影响尚不清楚。在 MN9D 多巴胺能神经元细胞中，我们分别使用 let-7d 模拟物和抑制剂上调和下调 let-7d 的表达，使用细胞计数试剂盒评估细胞活力，以及 TUNEL 染色测定和流式细胞术检查细胞死亡率，我们发现 let-7d 可以负调控 6-OHDA 诱导的细胞损伤。然后，我们通过双荧光素酶报告系统验证了 caspase-3 是 let-7d 的靶基因。此外，使用 caspase-3 siRNA 和 caspase-3 过表达载体（无 3'UTR）分别抑制和增加 caspase-3 的表达，我们发现 caspase-3 siRNA 可以逆转 let-7d 抑制剂诱导的细胞损伤，而过表达 caspase-3 可以逆转 let-7d 模拟物对 6-OHDA 诱导的细胞损伤的保护作用。综上所述，这些发现强烈表明 let-7d 在 DA 神经元细胞损伤中发挥重要作用，let-7d 的作用至少部分是通过抑制 caspase-3 的表达。
